While the United States food supply is among the safest in the world it is vulnerable to terrorist attacks as well as occasional accidental contamination from inadequate sanitation at packing and manufacturing facilities;e.g. the recent E. coli contamination of pre-washed bagged spinach. The research to be completed in this project will determine the feasibility of using Investigen's smartDNA" nucleic acid detection method to detect priority agents that have been intentionally added to a food. This is NOT intended as a food safety assay. It is intended as an assay for food security. Thus, the target organisms are biothreat agents as opposed to typical foodborne pathogens. The smartDNA detection system is an easy, rapid, isothermal (including room temperature), sequence-specific, colorimetric system for the detection of nucleic acids. Investigen currently has a smartDNA-based assay for the detection of Mycobacterium tuberculosis from sputum. On a separate project Investigen is developing prototypes of smartDNA assays and instrumentation for use by first responders for the detection of multiple bio-terrorism agents (Bacillus anthracis, Francisella tularensis and Yersinia pestis). This application builds on the results of these two initiatives and will determine the feasibility of using Investigen's smartDNA assay for the detection of nucleic acids from F. tularensis in food matrices. Detection of bacteria in foods is often confounded by the food matrix. Food matrices are complex, heterogeneous and not as well studied as clinical matrices. Detection systems that work in other matrices often are not able to detect the same bacteria when present in a food matrix. Thus Investigen will test the impact of food matrices on the smartDNA-based detection of F. tularensis. All aspects of the system will be tested, including the effect of food matrices (including processed samples from food matrices) on: (1) disruption of F. tularensis cells, (2) smartDNA-based target capture and release, and (3) smartDNA-based homogeneous detection of F. tularensis following sample processing. The results of these tests will be compared to the results of Investigen's standard smartDNA- based tests for the detection of F. tularensis. Based on the results, minimal inhibitory concentration each food matrix and the limit of detection of F. tularensis (isolated NA or whole cells - depending on the experiment) will be determined. Additionally, with a limited number of samples the assay's sensitivity and specificity will be evaluated. If the food matrices have a large negative effect on the smartDNA detection assay, then the buffers and reagents will be adjusted to attempt to mitigate this effect. The detection of Y. pestis and B. anthracis in food matrices will be a Phase II objective. PUBLIC HEALTH RELEVANCE: While the U.S. food supply is among the safest in the world, it is vulnerable to terrorist attacks and accidental contamination from inadequate sanitation at packing and manufacturing facilities. The research to be completed in this project will determine the feasibility of using Investigen's smartDNA nucleic acid detection technology to detect priority agents in a food matrix;a system that can be used in the field, in labs, at the point of harvest or manufacture and at importing locations such as U.S. borders and maritime ports. The rapid field deployable detection could aid in the identification and containment of contaminated food and reduce the impact on public health.